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随着生物化学与分子生物学理论的不断发展,基因治疗已成为医学界最活跃的研究领域之一。基因治疗载体在基因治疗过程中起一个转运和保护的作用。是基因治疗成功与否的关键。因此,基因治疗载体的开发对基因治疗的发展尤为重要,其有效开发将推动基因治疗向常规治疗方法的转变。目前,用于临床的基因载体大致分为病毒载体和非病毒栽体两类,它们都具有各自鲜明的优缺点,在临床治疗上亟需改进。脂质体在所有已用于临床试验的基因载体中仅次于病毒栽体,居第二位,是最有发展前景的非病毒载体。其中,阳离子脂质体由于具有对阴离子型聚电解质敏感,对带负电荷的DNA有较高的转运能力,还能转运RNA、核糖体及其他大电荷的分子和大分子物质进入细胞等优点,其转染效率比其他脂质体高出许多倍,因而被广泛应用于基因转移技术中。在阳离子类脂的结构中,连接键是一个非常重要的组成部分,它直接决定了阳离子脂质体的化学稳定性及被生物降解的能力,进而影响阳离子类脂的转染效率和细胞毒性。现在已合成出来的用于基因治疗的阳离子脂质体的连接键主要有醚键(如DOTMA、DMRIE)和酯键(如DOTAP)两种。前者由于过于稳定,在完成转染任务后不易分解,遗留在细胞内部对细胞产生毒害;而后者由于不够稳定,在转染过程中就易分解,对细胞的转染效率产生不利影响。因此,本课题组设计了以氨基甲酸酯键作为连接键的阳离子类脂,通过改变类脂前体分子中碳链长度、类脂分子合成中卤代烷碳链长度和卤素的类型来得到一系列的阳离子类脂,计划利用其pH敏感性和细胞内外的pH值差别,使在细胞外稳定的阳离子类脂在进入细胞内完成转染任务后可以分解成小分子有机物顺利排出,从而达到较好的转染效率和较低的细胞毒性。
With the continuous development of biochemistry and molecular biology theory, gene therapy has become one of the most active research fields in the medical field. Gene therapy vectors play a role in transport and protection during gene therapy. Gene therapy is the key to success. Therefore, the development of gene therapy vectors is particularly important for the development of gene therapy, and its effective development will promote the transformation from gene therapy to routine treatment. Currently, gene vectors for clinical use are broadly classified into two types: viral vectors and non-viral vectors. Both of them have their own distinctive advantages and disadvantages, which are in urgent need of improvement in clinical treatment. Liposomes are second only to viral vectors in all gene carriers that have been used in clinical trials, ranking second, and are the most promising non-viral vectors. Among them, the cationic liposomes have the advantages of being sensitive to anionic polyelectrolytes, being able to transport negatively charged DNA, transporting RNA, ribosomes and other large charged molecules and macromolecules into the cells, etc., Its transfection efficiency many times higher than other liposomes, which has been widely used in gene transfer technology. In the structure of cationic lipids, the linker is a very important part, which directly determines the cationic liposomes chemical stability and biodegradability, thus affecting the cationic lipid transfection efficiency and cytotoxicity. Cationic liposomes that have been synthesized for gene therapy are now mainly composed of ether linkages (eg DOTMA, DMRIE) and ester linkages (eg DOTAP). Due to the former is too stable, it is not easy to decompose after the completion of the transfection task, leaving the cells inside the cell toxic; the latter due to not stable enough, in the transfection process is easy to decompose, adversely affecting the transfection efficiency of the cells. Therefore, our group designed a series of cationic lipids with urethane bond as the linker, which can be obtained by changing the length of carbon chain in the precursor of lipids, the length of halogenated carbon chain in the lipid molecule synthesis and the type of halogen Of the cationic lipid, the planned use of its pH sensitivity and intracellular pH difference between the outside, so that the extracellular stable cationic lipids into the cells to complete the transfection task can be decomposed into small molecules of organic matter smoothly discharged, so as to achieve better Transfection efficiency and lower cytotoxicity.